TCA (Tricarboxylic Acid/ Krebs Cycle/ Citric Acid Cycle

The TCA (Tricarboxylic Acid) Cycle, also known as the Citric Acid Cycle or Krebs Cycle , is a crucial metabolic pathway that plays a significant role in cellular respiration. Here's a detailed description of the cycle:

- **Introduction**: The TCA Cycle is named after Hans Krebs, who first identified it. It's a series of chemical reactions used by all aerobic organisms to generate energy through the oxidation of acetate—derived from carbohydrates, fats, and proteins—into carbon dioxide.

- **Location**: In eukaryotic cells, the TCA Cycle occurs in the mitochondrial matrix, whereas, in prokaryotic cells, it takes place in the cytoplasm.

- **Process Overview**: The cycle starts with the combination of acetyl-CoA with oxaloacetate to form citrate. Through a series of eight steps, the cycle completes with the regeneration of oxaloacetate. Each turn of the cycle yields two molecules of carbon dioxide, three molecules of NADH, one molecule of FADH2, and one molecule of ATP (or GTP).

- **Steps of the Cycle**:
1. **Formation of Citrate**: Acetyl-CoA combines with oxaloacetate.
2. **Formation of Isocitrate**: Citrate is rearranged into isocitrate.
3. **Oxidation of Isocitrate**: Isocitrate is oxidized to α-ketoglutarate, producing NADH.
4. **Oxidation of α-Ketoglutarate**: α-Ketoglutarate is oxidized to succinyl-CoA, producing another NADH.
5. **Conversion of Succinyl-CoA to Succinate**: Energy from succinyl-CoA is used to form GTP (or ATP).
6. **Oxidation of Succinate**: Succinate is oxidized to fumarate.
7. **Hydration of Fumarate**: Fumarate is hydrated to malate.
8. **Oxidation of Malate**: Malate is oxidized to oxaloacetate, producing the final NADH.

- **Significance**: The TCA Cycle is not only pivotal for energy production but also provides intermediates for the synthesis of various biomolecules. It's tightly regulated and interconnected with other metabolic pathways

- **Regulation**: Key enzymes in the TCA Cycle are regulated by the energy needs of the cell, ensuring balance and efficiency in energy production

1. Aerobic Metabolism of
carbohydrate

2. The three stagesofrespiration
•Stage IAll the fuel molecules are oxidized to
acetyl-CoA.
•Stage IIThe acetyl-CoA is completely oxidized
into CO
2, electrons are collected by NAD and FAD
via the citric acid cycle.
•Stage IIIPassage of electrons through the electron
transportsystem to yield ATPfrom oxidative
phosphorylation.

3. Pyruvate + CoA + NAD+
acetylCoA + CO
2+ NADH + H
+
H
3CCCO

OO
CS
O
H
3C CoA
HSCoA
NAD
+
NADH
+ CO
2
Pyruvate Dehydrogenase
pyruvate acetyl-CoA
Pyruvate Oxidation

4. Pyruvate Dehydrogenaseis a large complex:
pyruvate dehydrogenase (E1),
dihydrolipoyl transacetylase (E2),
dihydrolipoyl dehydrogenase (E3)
Requires 5 coenzymes:
TPP, Lipoic Acid, Coenzyme A, FAD, NAD
+

5. Reactions of the PDH complex

6. Regulation of Pyruvate Dehydrogenase
1). Product inhibitionbyNADH &acetyl CoA
2). Covalent modification

7. Citric Acid Cycle
•Hans Krebs proposed the “citric acid cycle”
for the complete oxidation of pyruvate in
animal tissues in 1937 (1953 Nobel Prize
laureate).
•The tricaboxylic acid (TCA) Cycle, Krebs
Cycle

8. Citric Acid Cycle
•The common pathway leading to complete
oxidation of carbohydrates, fatty acids, and
amino acids to CO
2.
•Some ATP is produced, More NADH is
made ,NADH goes on to make more ATP in
electron transport and oxidative
phosphorylation
•A pathway providing many precursors for
biosynthesis

9. citric acid cycle overview CoASH
NADH
oxaloacetate citrate
malate
isocitrate
H
2O
CO
2
fumarate
ketoglutarate
(oxo-glutarate)
CoASHCoASH
FAD
FADH
GTP
succinate succinyl CoA
acetyl CoA
2C
6C4C
6C
5C
CO
2
4C4C
4C
4C
NADH
NADH
cis-aconitate
NAD
+
NAD
+
NAD
+
GDP, P
i

10. Individual reaction

11. COOH
C
CH
2
COOH
O
COOH
CH
2
C
CH
2
COOHHO
COOH
CH
3
C
SCoA
O
CoASH
citrate synthase
oxaloacetate
citrate
acetyl CoA Reaction 1: Citrate Synthase
A thioester: so a high
energy compound
Hydrolysis helps drive this
reaction forward
Allosteric,
-ATP,NADH,
succinyl-CoA

12. Fluoroacetate blocks the cycle
•Fluoroacetateis poisonous because it can
convert to fluorocitrate which is an
inhibitors of TCA cycle.

13. Reaction 2:
Aconitase
Dehydration followed
by hydrationcis-aconitate
aconitase
aconitase
isocitrate
citrate
H
H
COOH
CH
2
C
C
COOH
COOH
OH
COOH
CH
2
C
CH
2
COOHHO
COOH

14. Reaction 3:
Isocitrate
Dehydrogenase
FirstOxidative
decarboxylation
Allosteric enzyme
-ATP,NADHNAD
+
ketoglutarate
(oxo-glutarate)
isocitrate dehydrogenase
NADH
CO
2
isocitrate
COOH
CH
2
CH
2
C
COOH
O
H
H
COOH
CH
2
C
C
COOH
COOH
OH

15. Reaction 4:
-Ketoglutarate
Dehydrogenase
SecondOxidative
decarboxylationNAD
+
ketoglutarate dehydrogenase
succinyl CoA
CoASH
NADH
CO
2
ketoglutarate
(oxo-glutarate)
COOH
CH
2
CH
2
C
SCoA
O
COOH
CH
2
CH
2
C
COOH
O

16. Ketoglutarate dehydrogenase complex is very
similar to the pyruvate degydrogenase complex.
allosteric inhibitor: NADH,succinylCoA,ATP
•Five coenzymes used –
-TPP, CoASH, Lipoic acid,NAD
+
, FAD

17. GDP, P
i
succinyl CoA synthetase
succinyl CoA
succinate
GTP
CoASH
COOH
CH
2
CH
2
COOH
COOH
CH
2
CH
2
C
SCoA
O Reaction 5: Succinate thiokinase
Hydrolysis
(also called succinyl CoA synthetase)
A thioester: so a
high energy cpd
Substrate Level
Phosphorylation

18. Reaction 6:
Succinate
Dehydrogenase
Oxidation
e-carrier is FADsuccinate dehydrogenase
succinate
FADH
FAD
fumarate
COOH
CH
2
CH
2
COOH
COOH
CH
CH
COOH
2

19. Succinate Dehydrogenase
•Part of electron transport chain in the inner
membrane of mitochondria.
•Removal of H across a C-C bond is not
sufficiently exergonic to reduce NAD
+
,but it
does yield enough energy to reduce FAD.
•Malonate is a competitive inhibitor

20. Reaction 7:
Fumarase
Hydration
trans-addition of the elements
of water across the double bond,
forms L-malatefumarate
fumarase
H
2O
malate
COOH
CH
CH
COOH
COOH
CHOH
CH
2
COOH

21. Reaction 8: Malate Dehydrogenase
Oxidation
+NAD
+
malate
oxaloacetate
NADH
malate
dehydrogenase
COOH
C
CH
2
COOH
O
COOH
CHOH
CH
2
COOH

22. TCA Cycle SummaryCoASH
NADH
oxaloacetate citrate
malate
isocitrate
H
2O
CO
2
fumarate
ketoglutarate
(oxo-glutarate)
CoASHCoASH
FAD
FADH
GTP
succinate succinyl CoA
acetyl CoA
2C
6C4C
6C
5C
CO
2
4C4C
4C
4C
NADH
NADH
cis-aconitate
NAD
+
NAD
+
NAD
+
GDP, P
i
1 acetate
through the
cycle produces
2 CO
2, 1 GTP,
3NADH,
1FADH
2

23. Aerobic Nature of the Cycle
NADH and FADH
2must be reoxidized by the
electron transport chain.
Succinate Dehydrogenase is part of electron
transport chain in the inner membrane of
mitochondria.

24. Energetics
•Energy is conserved in the reduced
coenzymes NADH, FADH
2and one GTP
•NADH, FADH
2can be oxidized to
produce ATP by oxidative
phosphorylationacetyl CoA
TCA
3 NAD
+
3 NADH 9 ADP + 9 Pi
9 ATP
ETS
FADH
2
FAD
2 ADP + 2 Pi
2 ATP
ETS
1.5 1.5
1.5
7.5
7.5 7.5

25. ATP generated by the cycle
3 NADH 3 NAD
+
ETS
3*2.5=7.5 ATP
FADH
2 FAD
1.5 ATP
Substrate level
phosphorylation
1 GTP
10 ATP
Equivalents
Total
ETS

26. Glucose
glycolysis
2ATP(Substrate-level phosphorylation)2
2NADH( oxphos) 3-5
2Pyruvate
oxidative decarboxylation
2 NADH ( oxphos) 5
2 Acetyl CoA
TCA cycle 20
6 NADH
2 FADH
2
2 GTP
total 30-32ATP
25ATP
5-7ATP
CO2
ATP generated by complete oxidation of glucose

27. Regulation of the TCA Cycle
Again, 3 irreversible reactions are the key sites
•Citrate synthase-regulated by availability of
substrates -acetyl-CoA and oxaloacetate, citrate
is a competitive inhibitor;
Allosteric: -NADH , ATP,succinyl-CoA
•Isocitrate dehydrogenase–NADH,ATP inhibit,
ADP and NAD
+
Ca
++
activate
-Ketoglutarate dehydrogenase-NADH and
succinyl-CoA inhibit, AMP Ca
++
activate

28. Major regulatory
sites are irreversible
reactions

29. Anaplerotic reactions
•Anaplerotic(filling up) reactions replenish
citric acid cycle intermediates
•Amphibolic Nature of TCA Cycle means it
both Anabolic and Catabolic. TCA cycle
provides several of Intermediates for
Biosynthesis

31. Anaplerotic reactions
•PEP carboxylase-converts PEP to
oxaloacetate , Anaplerotic reaction in plants
and bacteria
•Pyruvate carboxylase-converts pyruvate to
oxaloacetate, a major anaplerotic reaction
in mammaliantissues
•Malic enzymeconverts pyruvate into malate

33. The Glyoxylate Cycle
An Anabolic Variant of the Citric Acid Cycle
for plants and bacteria
•Acetate-based growth -net synthesis of
carbohydrates and other intermediates from
acetate -is not possible with TCA
•Glyoxylate cycle offers a solution for plants and
some bacteria and algae
•The CO
2-evolving steps are bypassed and an
extra acetate is utilized
•Isocitrate lyase and malate synthase are the
short-circuiting enzymes

34. Discovery of Glyoxylate Cycle
Working together, Beevers and Kornberg showed
that malate synthase and isocitrate lyase, the two
enzymes that characterize the glyoxylate cycle, were
present in the endosperm of castor beans-1957

35. The glyoxylate cycle is particularly important
in species which synthesize carbohydrates
from two-carbon substrates, such as ethanol or
acetate,
and ,
In germinating plant seeds which must
synthesize their carbohydrates from stored
triacylglycerols.

36. Citrate
Isocitrate
Glyoxalate
Succinate
Oxaloacetate
Malate
Acetyl-CoA
Acetyl-CoA
Malate
sythase
Isocitrate
lyase

38. The Glyoxylate Cycle (Steps involved, Significance
and Regulation)
1. Condensation of acetyl-CoA with oxaloacetate to form
citrate.
2. Isomerization of citrate to isocitrate.
3. Cleavage of isocitrate to form succinate and glyoxylate:
4. Condensation of glyoxylate with acetyl-CoA to
yield malate.
5. Oxidation of malate to form oxaloacetate.Malate is
oxidized subsequently to oxaloacetate by the
enzymemalate dehydrogenase.
6. Oxaloacetate then starts another turn of the cycle.

39. Similarities with TCA cycle:
The glyoxylate cycle uses five of the eight enzymes associated with
thetricarboxylic acid cycle:
citrate synthase,aconitase,succinate dehydrogenase,fumarase,
andmalate dehydrogenase.
The two cycles differ in that in the glyoxylate cycle,isocitrateis
converted intoglyoxylateandsuccinateby isocitrate lyase (ICL)
instead of into α-ketoglutarate.
This bypasses the decarboxylation steps that take place in the citric
acid cycle (TCA cycle), allowing simple carbon compounds to be
used in the later synthesis of macromolecules, including glucose.
Glyoxylateis subsequently combined withacetyl-CoAto
producemalate, catalyzed by malate synthase.Malate is also formed
in parallel from succinate by the action of succinate dehydrogenase
and fumarase.

40. Significanceofglyoxylatecycle:
ItisabypassreactionofTCAcycle.
Oxaloacetateformedinthispathwaycanbeusedtosynthesize
glucoseviagluconeogenesis.
Itoccursinbacteriawhentheyareculturedinacetaterichcarbon
source(acetatebeingthesolesourceofcarbon).
Thispathwayisverysignificantingerminatingseedswherethe
storedtriacylglycerols(fat)isconvertedintosugarstomeetthe
energyneeds.
Whenhigherfattyacidsareoxidizedintoacetyl-CoAwithout
formingpyruvates,thenacetyl-CoAentersintoglyoxylatecycle.

41. Coordinatedregulationof
GlyoxylateandTCAcycles:
Regulationofisocitratedehydrogenase
activitydeterminesthepartitioningof
isocitratebetweentheglyoxylateand
citricacidcycles.
Whentheenzymeisinactivatedby
phosphorylation(byaspecificprotein
kinase),isocitrateintobiosynthetic
reactionsviaglyoxylatecycle.
Whentheenzymeisactivatedby
dephosphorylation(byaspecific
phosphatase),isocitrateenterstheTCA
cycleandATPisproduced.